The goal of this proposal is to understand molecular mechanisms regulating TRP, the archetypal member of a new class of proteins referred to as store-operated channels (SOC). SOCs are activated by Ca2+ release from internal stores and appear to be responsible for a prevalent Ca2+ influx pathway. Vertebrate SOCs were recently identified, but mechanisms that regulate SOC's in general are poorly understood. Three general strategies to study SOCs are to be used. The first will employ in vitro binding to characterize the nature of the protein interactions in the TRP signaling complex. The second is to probe TRP function in vivo using the Drosophila visual system. A series of site-specific mutations will be made in TRP and the effects of the mutations will be assessed following germ-line transformation of the mutant genes. The third strategy is an electrophysiological analysis of members of the TRPC family expressed in cultured cells. The specific aims of the proposal focus on several hypotheses: (1) that INAD binds directly to multiple proteins required in vision to facilitate feedback regulation; (2) that CaM functions in feedback regulation of TRP; (3) that TRP is regulated by interaction with GTP; (4) that the vertebrate TRPC proteins form heteromultimers; and (5) that TRPR1 and TRPR2 define a new subfamily of TRP-like channels.